title: Characterization of human myoblast differentiation for tissue-engineering purposes by quantitative gene expression analysis.
abstract: Tissue engineering of skeletal muscle is an encouraging possibility for the treatment of muscle loss through the creation of functional muscle tissue in vitro from human stem cells. Currently, the preferred stem cells are primary, non-immunogenic satellite cells ( = myoblasts). The objective of this study was to determine the expression patterns of myogenic markers within the human satellite cell population during their differentiation into multinucleated myotubes for an accurate characterization of stem cell behaviour. Satellite cells were incubated (for 1, 4, 8, 12 or 16 days) with a culture medium containing either a low [ = differentiation medium (DM)] or high [ = growth medium (GM)] concentration of growth factors. Furthermore, we performed a quantitative gene expression analysis of well-defined differentiation makers: myogenic factor 5 (MYF5), myogenin (MYOG), skeletal muscle αactin1 (ACTA1), embryonic (MYH3), perinatal (MYH8) and adult skeletal muscle myosin heavy chain (MYH1). Additionally, the fusion indices of forming myotubes of MYH1, MYH8 and ACTA1 were calculated. We show that satellite cells incubated with DM expressed multiple characteriztic features of mature skeletal muscles, verified by time-dependent upregulation of MYOG, MYH1, MYH3, MYH8 and ACTA1. However, satellite cells incubated with GM did not reveal all morphological aspects of muscle differentiation. Immunocytochemical investigations with antibodies directed against the differentiation markers showed correlations between the gene expression and differentiation. Our data provide information about time-dependent gene expression of differentiation markers in human satellite cells, which can be used for maturation analyses in skeletal muscle tissue-engineering applications.
authors.affiliation: Department of Otolaryngology, University of Heidelberg, Mannheim, Germany. jens.stern-straeter@umm.de
authors.forename: Jens
authors.initials: J
authors.lastname: Stern-Straeter
authors.forename: Gabriel Alejandro
authors.initials: GA
authors.lastname: Bonaterra
authors.forename: Stefan S
authors.initials: SS
authors.lastname: Kassner
authors.forename: Stefanie
authors.initials: S
authors.lastname: Zügel
authors.forename: Karl
authors.initials: K
authors.lastname: Hörmann
authors.forename: Ralf
authors.initials: R
authors.lastname: Kinscherf
authors.forename: Ulrich Reinhart
authors.initials: UR
authors.lastname: Goessler
mesh_terms: D015415:Biomarkers / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics*; D002459:Cell Fusion; D049109:Cell Proliferation / Q000187:drug effects; D003470:Culture Media / Q000494:pharmacology; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007150:Immunohistochemistry; D008875:Middle Aged; D032446:Myoblasts / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D009928:Organ Specificity / Q000187:drug effects; D012313:RNA / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D032496:Satellite Cells, Skeletal Muscle / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D023822:Tissue Engineering / Q000379:methods*
chemical_list: D015415:Biomarkers; D003470:Culture Media; D012313:RNA